Cephalosporins having an α-acylaminoacetic acid side chain

ABSTRACT

Compounds of the formula ##STR1## wherein R 1  denotes optionally substituted phenyl, thienyl, furyl or 1,4-cyclohexadienyl, R 2  represents a free carboxyl group or an esterified carboxyl group which can be split physiologically, R 3  represents hydrogen, lower alkoxy or an optionally substituted methyl group and B represents an optionally substituted six-membered ring with 1 to 3 ring nitrogen atoms, which is bonded to the carbonyl group --C(═O)-- by one of its carbon atoms, the nitrogen atoms of a monocyclic six-membered ring having 2 nitrogen atoms being either adjacent or separated by two ring carbon atoms, and the salts of such compounds which have a salt-forming group, including the inner salts, for example the 7β-[D(-)-α-(3,5-Dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanic acid, have antibiotic activity.

This is a division of application Ser. No. 576,398 filed May 9, 1975,now U.S. Pat. No. 4,041,161.

The invention relates to new therapeutically valuable derivatives of7-amino-ceph-3-em-4-carboxylic acid and their salts, processes for theirmanufacture and pharmaceutical preparations which contain the newcompounds.

In particular, the invention relates to compounds of the formula##STR2## wherein R₁ denotes optionally substituted phenyl, thienyl,furyl or 1,4-cyclohexadienyl, R₂ represents a free carboxyl group or anesterified carboxyl group which can be split physiologically, R₃represents hydrogen, lower alkoxy or an optionally substituted methylgroup and B represents an optionally substituted six-membered ring with1 to 3 ring nitrogen atoms, which is bonded to the carbonyl group-C(═O)-by one of its carbon atoms, the nitrogen atoms of a monocyclicsix-membered ring having 2 nitrogen atoms being either adjacent orseparated by two ring carbon atoms, and the salts of such compoundswhich have a salt-forming group, including the inner salts.

Unless defined otherwise, the general expressions employed in thepreceding and following text have the following meanings:

Lower alkyl is a straight-chain or branched alkyl group with 1 to 7,preferably with up to 4, carbon atoms, and denotes, for example, methyl,ethyl, propyl, isopropyl, butyl, isobutyl, tert.-butyl, pentyl, hexyl orheptyl.

Lower alkoxy is, for example, methoxy, ethoxy, propoxy, isopropoxy,butoxy, isobutoxy, tert.-butoxy, pentyloxy, hexyloxy or heptyloxy andlower alkylmercapto is, for example, methylmercapto, ethylmercapto,propylmercapto, isopropylmercapto, butylmercapto, isobutylmercapto,tert.-butylmercapto, pentylmercapto, hexylmercapto or heptylmercapto.

Lower alkanoyl is a straight-chain or branched lower alkylcarbonyl groupwith 1 to 8, preferably up to 5, carbon atoms, and denotes, for example,formyl, acetyl, propionyl, butyryl, valeryl, isovaleryl orheptanecarbonyl.

Halogen is fluorine, chlorine or bromine.

Aryl is a monocyclic or polycyclic, such as bicyclic or tricyclic,aromatic radical with up to 14 carbon atoms and is, for example, phenyl,naphthyl or anthranyl, which can optionally be substituted, for exampleby hydroxyl, lower alkoxy, lower alkyl, halogen or nitro. Aroyl is acorresponding arylcarbonyl radical, for example benzoyl.

Examples of substituents of the phenyl group R₁ are optionally protectedhydroxyl, lower alkyl, such as methyl, lower alkoxy, such as methoxy,halogen atoms, such as fluorine or chlorine, halogeno-lower alkyl, suchas trifluoromethyl, optionally protected amino, nitro, optionallyprotected amino-lower alkyl, such as aminomethyl, carbamoyl,carbamoyloxy or carbamoylamino, which are optionally N-monosubstitutedor N,N-disubstituted, for example by lower alkyl, such as methyl, oracyl, especially lower alkanoyl, such as acetyl, the substituents beingin the o-, m- or preferably the p-position.

The substituent R₂ is, in particular, a free carboxyl group or anesterified carboxyl group which can be split physiologically, forexample an esterified carboxyl group which can be split enzymatically.

Esterified carboxyl groups which can be split physiologically are, aboveall, those which can be split enzymatically or be split by the acidgastric juice. These esters are readily resorbable in the organism andcan therefore be used therapeutically as such. Esters of this type aredescribed, for example, in British Patent specification No. 1,229,453,in Belgian Pat. No. 789,821 and in German Patent Applications DT1,951,012, DT 2,228,012 and DT 2,230,620. Such esters are derived, forexample, from indan-1-ol or 5-hydroxyindan-1-ol or3,4-benzo-5-oxo-tetrahydro-2-furanol or from alcohols of the formulaHO-CH₂ OCO-R₂ ', wherein R₂ ' can represent an alkyl radical oraminoalkyl radical or a cycloalkyl radical with 3-7 carbon atoms. Inparticular, R₂ ' denotes a lower alkyl radical, such as methyl, ethyl,isopropyl and above all tert.-butyl, an α-amino-lower alkyl radical,such as 1-amino-2-methylpropyl or 1-amino-3-methylbutyl, or acyclopentyl or cyclohexyl radical.

A lower alkoxy group R₃ contains 1 to 7, preferably 1 to 4, carbon atomsand is in particular methoxy (compare Netherlands Application No.73/09,136). Substituents of the methyl group R₃ are, above all, free,esterified or etherified hydroxyl or mercapto groups, and alsooptionally N-substituted carbamoyloxy or thiocarbamoylmercapto groups,or quaternary ammonium groups.

An esterified hydroxyl or mercapto group in a substituted methyl groupR₃ contains, as the acid radical, above all the radical of a carboxylicacid or thiocarboxylic acid, for example lower alkanoyl, which isoptionally substituted by halogen atoms, especially chlorine, such asformyl, propionyl, butyryl, pivaloyl or chloroacetyl, but especiallyacetyl, or aroyl or aryl-lower alkanoyl optionally substituted by, forexample, lower alkyl, lower alkoxy, halogen or nitro, for examplebenzoyl or phenylacetyl, and also, as a thiocarboxylic acid radical, inparticular thioaroyl which is optionally substituted as mentioned, andabove all thiobenzoyl. Esterified mercapto groups can in particular alsocontain heteroyl, wherein the heterocyclyl radical preferably contains5-6 ring members and, as hetero-atoms, nitrogen, optionally in theN-oxidised form, and/or oxygen or sulphur, for example optionally1-oxidised pyridyl, pyrimidyl, pyridazinyl, thiadiazolyl, oxadiazolyl orN-methyltetrazolyl. In addition, hydroxyl groups esterified by hydrogenhalide acids should be mentioned; the methyl group R₅ can therefore besubstituted by, for example, fluorine, chlorine or bromine.

Etherified hydroxyl groups in a substituted methyl group R₃ aredescribed, for example, in Belgian Pat. No. 719,710. Lower alkoxy, suchas methoxy, ethoxy or n-propoxy, should be singled out.

Etherified mercapto groups in a substituted methyl group R₃ for examplecontain, as the etherifying radicals, lower alkyl, for example methyl,and also optionally substituted phenyl or heterocyclyl. Phenyl can besubstituted by, for example, lower alkyl, lower alkoxy, halogen ornitro. The heterocyclyl radicals preferably have 5-6 ring members andcontain, as hetero-atoms, nitrogen, optionally in the N-oxidised form,and/or oxygen or sulphur. Examples to be mentioned are optionally1-oxidised pyridyl, pyrimidyl, pyridazinyl, pyrazinyl, imidazolyl,imidazolidyl, purinyl, triazolyl and tetrazolyl. These radicals can besubstituted, for example, by lower alkyl, lower alkoxy, hydroxyl orhalogen. Optionally substituted heterocyclyl radicals of aromaticcharacter, with 5 ring atoms which comprise 2 nitrogen atoms and afurther oxygen or sulphur atom or 1 to 2 further nitrogen atoms, shouldbe singled out particularly. Preferred substituents are lower alkylradicals with 1-5 carbon atoms such as methyl, ethyl, propyl, isopropyl,n-butyl, isobutyl and tert.butyl, lower alkoxy or lower alkylthioradicals with 1-5 carbon atoms, particularly methylthio, cycloalkylradicals with 3-7 carbon atoms, for example cyclopentyl or cyclohexyl,or aryl radicals, such as phenyl or substituted phenyl, for examplephenyl substituted by one or more nitro groups or halogen atoms or loweralkyl or lower alkoxy groups, or unsubstituted or substituted thienyl,particularly 2-thienyl, or thienyl substituted as indicated for phenyl,or, in particular, optionally monosubstituted or disubstituted aminogroups, for example amino, lower alkylamino, which optionally containscarboxyl or amino, such as methylamino, 2-carboxyethylamino or2-aminoethylamino, acylamino, such as lower alkanoylamino, whichoptionally contains carboxyl or amino, such as acetylamino or especiallyβ-carboxypropionylamino or α-aminoacetylamino, or loweralkoxycarbonylamino, such as tert.-butoxycarbonylamino ortert.-amyloxycarbonylamino, or sulphonylamino.

The following should be mentioned as examples of the heterocyclylradical which etherifies the mercapto group:

Triazolyl optionally substituted by lower alkyl and/or aryl, such asphenyl, such as 1H-1,2,3-triazol-5-yl, 1-methyl-1H-1,2,3-triazol-4-yl,1H-1,2,4-triazol-3-yl, 5-methyl-1H-1,2,4-triazol-3-yl,3-methyl-1-phenyl-1H-1,2,4-triazol-5-yl,4,5-dimethyl-4H-1,2,4-triazol-3-yl and 4-phenyl-4H-1,2,4-triazol-3-yl,tetrazolyl optionally substituted by lower alkyl or aryl, such as phenylor chlorophenyl, such as 1H-tetrazol-5-yl, 1-methyl-1H-tetrazol-5-yl,1-ethyl-1H-tetrazol-5-yl, 1-n-propyl-1H-tetrazol-5-yl,1-phenyl-1H-tetrazol-5-yl and 1-p-chlorophenyl-1H-tetrazol-5-yl,thiazolyl optionally substituted by, for example, lower alkyl orheterocyclyl, such as thienyl, such as 2-thiazolyl,4-(2-thienyl)-2-thiazolyl and 4,5-dimethyl-2-thiazolyl, thiadiazolyloptionally substituted by lower alkyl, amino, lower alkylamino whichoptionally contains carboxyl or amino, such as methylamino,2-carboxyethylamino or 2-aminoethylamino, or lower alkanoylamino whichoptionally contains carboxyl or amino, such as, in particular,β-carboxypropionylamino or α-aminoacetylamino, such as1,3,4,-thiadiazol-2-yl, 2-methyl-1,3,4-thiadiazol-5-yl,2-ethyl-1,3,4-thiadiazol-5-yl, 2-n-propyl-1,3,4-thiadiazol-5-yl,2-isopropyl-1,3,4-thiadiazol-5-yl, 2-amino-1,3,4-thiadiazol-5-yl and2-(β-carboxypropionylamino)-1,3,4-thiadiazol-5-yl, 5-thiatriazolyl,oxazolyl, isoxazolyl or oxadiazolyl optionally substituted by, forexample, lower alkyl and/or aryl, such as 5-oxazolyl,4-methyl-5-oxazolyl, 2-oxazolyl, 4,5-diphenyl-2-oxazolyl,3-methyl-5-isoxazolyl, 1,2,4-oxadiazol-5-yl,2-methyl-1,3,4-oxadiazol-5-yl, 2-phenyl-1,3,4-oxadiazol-5-yl,5-β-nitrophenyl-1,3,4-oxadiazol-2-yl and2-(2-thienyl)-1,3,4-oxadiazol-5-yl, as well as bicyclic heterocyclylradicals which are optionally substituted by halogen or nitro, such as2-benzimidazolyl, 5-chloro-2-benzimidazolyl, 2-benzoxazolyl,5-nitro-2-benzoxazolyl, 5-chloro-2-benzoxazolyl, s-triazolo[4.3-a]pyrid-3-yl, 3H-v-triazolyl[4.5-b]pyrid-5-yl, purin-2-yl,purin-6-yl and 8-chloro-2-methylpurin-6-yl.

An optionally N-substituted carbamoyloxy group or thiocarbamoylmercaptogroup in a substituted methyl group R₃ is, for example, a group of theformula --O--CO--NH--R^(II) (French Pat. No. 1,463,831), wherein R^(II)is hydrogen or an optionally halogen-substituted lower alkyl radical, ora group of the formula --S--C(═S)--N(R^(II))(R^(III)), wherein R^(II)has the above-mentioned meaning and R^(III) represents hydrogen or hasthe abovementioned meaning of R^(II) [compare J. Med. Chem. 8, 174(1965)]. Above all, R^(II) is methyl, ethyl or chlorine-substitutedmethyl or ethyl, especially β-chloroethyl.

In a quaternary ammonium-methyl group R₃, the ammonium part ispreferably an unsubstituted or substituted pyridinium group. Examples ofsubstituents of the pyridinium group which should be mentioned are thoselisted in Antimicrobial Agents and Chemotherapy 1966, page 573-580, suchas unsubstituted or substituted, for example hydroxyl-substituted orcarboxyl-substituted, lower alkyl, for example methyl, ethyl, propyl,hydroxymethyl or carboxymethyl, halogen, such as fluorine, chlorine,bromine or iodine, or trifluoromethyl, hydroxyl, sulpho, carboxyl,cyano, lower alkoxycarbonyl, such as methoxycarbonyl or ethoxycarbonyl,lower alkylcarbonyl, such as methylcarbonyl, and, in particular,carbamoyl which is unsubstituted or substituted, for example carbamoylsubstituted by lower alkyl, hydroxy-lower alkyl or halogeno-lower alkyl,especially chloro-lower alkyl, such as N-methylcarbamoyl,N-isopropylcarbamoyl and N-β-chloroethylcarbamoyl, but above allcarbamoyl. The substituents can be in the 2-, 3-and/or 4-position, butare preferably in the 3- or 4-position.

The radical B can be monocyclic, bicyclic or tricyclic and in particularcomprises optionally polysubstituted and/or optionally partiallyhydrogenated pyridine, pyrazine, pyridazine, triazine, quinoline,isoquinoline, naphthyridine or pyrido-pyrimidine radicals which areoptionally substituted at the nitrogen-containing ring by 1 or 2optionally, in particular physiologically splittably, esterified, oretherified hydroxyl or mercapto groups or optionally mono- or di-loweralkylated or lower alkanoylated amino groups or by halogen, and thetautomers of these radicals.

Radicals B to be singled out for example have the formula (B₁) ##STR3##wherein R₄ denotes lower alkyl, especially methyl, and R₅ and R₆substitute adjacent ring carbon atoms and conjointly represent a loweralkylene radical, which optionally carries an oxo group, especially a1,3-propylene, 1,4-butylene or 1,5-pentylene radical, or represent aradical of the formula ##STR4## wherein the two R₇ independently of oneanother denote hydrogen, hydroxyl, lower alkyl, especially methyl, loweralkoxy, especially methoxy, lower alkylmercapto, especiallymethylmercapto, lower alkanoyl, especially acetyl, lower alkanoylamino,especially acetylamino, lower alkoxycarbonyloxy, especiallyethoxycarbonyloxy, lower alkylsulphonyl, especially methylsulphonyl, oraryl, especially phenyl, or the two R₇ conjointly with the group--CH═CH-- form a thiazole, isothiazole, pyrrole, furane or benzene ringwhich, for example, can be substituted by an oxo group, a lower alkylgroup, such as a methyl group, or a lower alkanoyl group, such as anacetyl group, and wherein the broken line denotes a 4,5- or 5,6-doublebond, or have the formula (B₂) ##STR5## wherein R₈ denotes a freehydroxyl or mercapto group, an etherified hydroxyl or mercapto group,such as a lower alkylated, for example methylated, hydroxyl or mercaptogroup, or an esterified hydroxyl or mercapto group, especially aphysiologically splittable esterified hydroxyl or mercapto group, suchas a hydroxyl or mercapto group esterified with a half-ester of carbonicacid, for example monoethyl carbonate, a free amino group or a mono- ordi-lower alkylated, such as methylated, amino group, or a loweralkanoylated, such as acetylated, or a carbamoylated amino group orhalogen, especially chlorine, R₉ represents hydrogen or has the meaningof R₈ or represents nitrile, lower alkyl, such as methyl, hydroxy-loweralkyl, such as 1-hydroxyethyl, lower alkanoyl, such as acetyl, aryl,especially phenyl, or arylcarbonyl, especially phenylcarbonyl, R₁₀represents hydrogen, nitrile, lower alkyl, especially methyl, loweralkanoyl, especially acetyl, or aryl, especially phenyl, or R₉ and R₁₀conjointly represent a lower alkylene radical, which optionally carriesan oxo group, especially a 1,3-propylene, 1,4-butylene or 1,5-pentyleneradical, or R₉ and R₁₀ substitute adjacent ring carbon atoms andconjointly represent a radical of the formula ##STR6## wherein the twoR₁₁ independently of one another denote hydrogen, hydroxyl, optionallyhalogenated lower alkyl, such as methyl or trifluoromethyl, loweralkoxy, such as ethoxy, lower alkanoyloxy, such as acetoxy, mercapto,lower alkylmercapto, especially methylmercapto, lower alkanoyl,especially acetyl, amino, mono- or di-lower alkylamino, such asdimethylamino, lower alkanoylamino, such as acetylamino, loweralkyloxycarbonylamino optionally substituted by phenyl, such asethoxycarbonylamino or benzyloxycarbonylamino, lower alkylsulphonyl,especially methylsulphonyl, or aryl, especially phenyl, or the two R₁₁conjointly with the group --CH═CH-- form a thiazole, isothiazole,pyrrole, furane or benzene ring, which can be substituted by an oxogroup, a lower alkyl group, such as a methyl group, or a lower alkanoylgroup, such as an acetyl group, or the two R₁₁ in adjacent positionsconjointly denote the methylenedioxy group or a lower alkylene radical,especially the 1,3-propylene, 1,4-butylene or 1,5-pentylene radical, orR₉ and R₁₀ conjointly represent a group of the formula ##STR7## whereinR₁₂ denotes hydrogen, hydroxyl or lower alkyl, especially methyl, loweralkoxy, especially methoxy, amino, mono- or di-lower alkylamino,especially dimethylamino, or lower alkoxycarbonylamino optionallysubstituted by phenyl, such as ethoxycarbonylamino orbenzyloxycarbonylamino, and R₁₃ denotes hydrogen, lower alkyl,especially methyl, or lower alkoxy, especially methoxy, or have theformula (B₃) ##STR8## wherein R₈ and R₉ have the meaning mentioned underformula (B₂), or have the formula (B₄) ##STR9## wherein R₈ and R₉ havethe meaning mentioned under formula (B₂), or have the formula (B₅)##STR10## wherein R₁₄ and R₁₅ independently of one another denotehydrogen, halogen, especially chlorine, a free hydroxyl or mercaptogroup, an etherified hydroxyl or mercapto group, especially a loweralkylated, such as methylated, hydroxyl or mercapto group, or anesterified hydroxyl or mercapto group, especially a physiologicallysplittable esterified hydroxyl or mercapto group, such as a hydroxyl ormercapto group esterified with a half-ester of carbonic acid, forexample monoethyl carbonate, or an optionally mono- or di-loweralkylated, especially dimethylated, amino group or a lower alkanoylated,such as acetylated, or a carbamoylated amino group, or have the formula(B₆) ##STR11## wherein R₁₄ and R₁₅ have the meaning mentioned underformula (B₅), the substituent R₁₄ preferably occupying the 5-position ofthe as-triazine ring, or, where appropriate, have a formula tautomericthereto.

Compounds to be singled out are those of the formula I, wherein R₁ hasthe meaning mentioned under formula I and phenyl is in particularsubstituted by hydroxyl, such as p-hydroxyl, R₂ denotes a carboxylgroup, R₃ denotes hydrogen, lower alkoxy, lower alkanoyloxymethyl, loweralkoxymethyl, aroylthiomethyl, optionally substituted pyridiniomethyl,or heterocyclylmercaptomethyl, the heterocyclyl radical having 5 ringatoms and being of aromatic character, being optionally substituted andcontaining, in addition to 2 nitrogen atoms, at least one further ringhetero-atom from the group nitrogen, oxygen or sulfur, especiallyoptionally substituted thiadiazolylmercaptomethyl, such asthiadiazolylmercaptomethyl substituted by amino or3-carboxypropionylamino, trizolylmercaptomethyl, ortetrazolylmercaptomethyl optionally substituted by lower alkyl, such asmethyl, and B denotes one of the groups B₁ to B₆, especially a group B₂,B₃ or B₆ which is mono- to di-substituted exclusively by hydroxyl,halogen, such as chlorine, mercapto and/or methylmercapto, and saltsthereof.

The invention relates above all to compounds of the formula I, whereinR₁ has the meaning mentioned under formula I and in particularrepresents phenyl, R₂ denotes a carboxyl group, R₃ denotes hydrogen,lower alkoxy, such as methoxy, lower alkanoyloxymethyl, such asacetoxymethyl, aroylthiomethyl, such as benzoylthiomethyl, optionallysubstituted pyridiniomethyl, such as 4-carbamoylpyridiniomethyl, orheterocyclylmercaptomethyl, the heterocyclyl radical having 5 ring atomsand being of aromatic character, being optionally substituted andcontaining, in addition to 2 nitrogen atoms, at least one further ringhereto atom from the group nitrogen, oxygen or sulphur, especiallyoptionally substituted thiadiazolylmercaptomethyl, such asthiadiazolylmercaptomethyl substituted by amino or3-carboxypropionylamino, triazolylmercaptomethyl, ortetrazolylmercaptomethyl which is optionally substituted by lower alkyl,such as methyl, such as 2-amino-1,3,4-thiadiazol-5-yl-mercaptomethyl,2-(3-carboxypropionylamino)-1,3,4-thiadiazol-5-ylmercaptomethyl,1,2,3-1H-triazol-5-ylmercaptomethyl or1-methyl-1H-tetrazol-5-ylmercaptomethyl, and the group B denotespyridine which is mono- or di-substituted by hydroxyl or chlorine,quinoline substituted by hydroxyl, pyridazine substituted by hydroxyl,or 1,3,4-triazine di-substituted by hydroxyl and/or methylmercapto, or atautomer thereof, and to salts thereof.

Compounds to be singled out in particular are those of the formula I,wherein R₁ has the meaning mentioned under formula I and in particularrepresents phenyl, R₂ denotes a carboxyl group, R₃ denotes lower alkoxy,such as methoxy, lower alkanoyloxymethyl, such as acetoxymethyl,aroylthiomethyl, such as benzoylthiomethyl, optionally substitutedpyridiniomethyl, such as 4-carbamoyl-pyridiniomethyl, orheterocyclylmercaptomethyl, the heterocyclyl radical having 5 ring atomsand being of aromatic character, being optionally substituted andcontaining, in addition to 2 nitrogen atoms, at least one further ringhetero-atom from the group nitrogen, oxygen or sulphur, especiallyoptionally substituted thiadiazolylmercaptomethyl,triazolylmercaptomethyl, or tetrazolylmercaptomethyl, optionallysubstituted by lower alkyl, such as methyl, such as2-amino-1,3,4-thiadiazol-5-ylmercaptomethyl,2-(3-carboxypropionylamino)-1,3,4-thiadiazol-5-ylmercaptomethyl,1,2,3-1H-triazol-5-ylmercaptomethyl or1-methyl-1H-tetrazol-5-ylthiomethyl, and the group B denotes2-hydroxypyrid-5-yl, 6-hydroxypyridazin-3-yl, 2-hydroxyquinolin-4-yl,2,6-dichloropyrid-4-yl, 3,5-dihydroxy-1,2,4-triazin-6-yl or3-methylthio-6-hydroxy-1,2,4-triazin-6-yl, or a tautomer thereof, andtheir salts.

Salts of compounds of the present invention are, above all,pharmaceutically usable non-toxic salts of those compounds which areable to form salts with bases. Such salts are, above all, metal salts orammonium salts, such as alkali metal salts or alkaline earth metalsalts, for example sodium, potassium, magnesium, calcium or aluminiumsalts, as well as ammonium salts with ammonia or suitable organicamines, the amines used for forming the salt being, above all,aliphatic, cycloaliphatic, cycloaliphatic-aliphatic and araliphaticprimary, secondary or tertiary monoamines, diamines or polyamines, aswell as heterocyclic bases, such as lower alkylamines, for exampletriethylamine, hydroxy-lower alkylamines, for example2-hydroxyethylamine, bis-(2-hydroxyethyl)-amine ortris-(2-hydroxyethyl)-amine, basic aminoacids such as lysine, ornithineand arginine, basic aliphatic esters of carboxylic acids, for example4-aminobenzoic acid 2-diethylaminoethyl ester, lower alkyleneamines, forexample 1-ethylpiperidine, cycloalkylamines, for examplebicyclohexylamine, or benzylamines, for exampleN,N'-dibenzyl-ethylenediamine, and also bases of the pyridine type, forexample pyridine, collidine or quinoline.

Further salts are derived from the compounds according to the inventionwhich have a basic group, for example an unsubstituted or monoalkylatedor dialkylated amino group. Such compounds either form inner salts withthe free carboxyl group R₂ or can be converted into a salt with apharmaceutically usable, non-toxic organic or inorganic acid. Suitableacids are organic carboxylic acids or sulphonic acids, for examplealkanoic acids, such as trifluoroacetic acid, or methanesulphonic acid,or aromatic acids, such as benzoic acid or benzenesulphonic acid, orinorganic acids, such as hydrochloric acid, sulphuric acid or phosphoricacid.

The new compounds can be in the form of mixtures of isomers, for exampleracemates, or in the form of individual isomers, for example opticallyactive antipodes.

The new compounds of the formula I exhibit a pharmacological action, inparticular an especially pronounced antibacterial action. Thus they areactive against Gram-positive bacteria, such as staphylococci, or againstGram-negative bacteria, such as enterobacteriaceae, for example strainsof Escherichia, and especially against strains of Pseudomonas.

For example, they inhibit the growth of enterobacteriaceae andstaphylococci and pseudomonads in vitro in concentrations of about 0.15to 60 μg/ml. In mice, they are active, on subcutaneous administration,against staphylococci, such as Staphylococcus aureus, in a dosage rangeof about 8 to about 100 mg/kg, against enterobacteriaceae, such asEscherichia coli, in a dosage range from about 250 to about 280 mg/kg,and against pseudomonads, such as Pseudomonas aeruginosa, in a dosagerange of about 50 to about 200 mg/kg. The compounds of the formula I ortheir pharmaceutically acceptable salts, for example7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid, can therefore be used to combat infections which are caused bysuch microorganisms, and also as feedingstuff additives, for thepreservation of foodstuffs or as disinfectants.

The compounds of the present invention are manufactured according tomethods which are in themselves known. For example, they can bemanufactured by

(a) acylating the primary amino group in a compound of the formula II##STR12## wherein R₁ has the indicated meaning, R₂ ^(o) has the meaningof R₂ or represents an easily splittable esterified carboxyl group andR₃ has the indicated meaning, or in a salt thereof, by treatment with anacid of the formula B-COOH (III), wherein B has the indicated meaning,or with a reactive functional derivative thereof, or by

(b) acylating a compound of the formula IV ##STR13## wherein R₂ ^(o) hasthe meaning of R₂ or represents an easily splittable esterified carboxylgroup and R₃ has the indicated meaning, or a salt thereof, with acarboxylic acid of the formula (V) ##STR14## wherein R₁ and B have theindicated meanings, or with a reactive functional derivative thereof,and, if appropriate, in a resulting compound, splitting off a protectivegroup present in the radical B and/or converting an easily splittableesterified carboxyl group R₂ ^(o) into a free carboxyl group R₂ or aphysiologically splittable esterified carboxyl group R₂ and/or, ifdesired, converting an optionally substituted methyl group R₃ intoanother group R₃ and/or, if desired, converting a compound obtained asthe free acid into a salt, or a salt obtained into the free acid, and/orseparating an isomer mixture obtained into the individual isomers.

An easily splittable esterified carboxyl group R₂ ^(o) in a startingmaterial of the formula II or IV is, in particular, an ester group whichcan be split to the free carboxyl group in a neutral, acid or weaklyalkaline medium, solvolytically, for example hydrolytically,alcoholytically or acidolytically, or reductively, for examplehydrogenolytically.

Esterified carboxyl groups R₂ ^(o) which can easily be split bysolvolysis with a solvent containing hydroxyl groups, for example wateror alcohols, such as, for example, methanol or ethanol, preferably underneutral conditions, are above all those which are derived from silylalcohol or stannyl alcohol. Such groups are described, for example, inBritish Patent specifications 1,073,530 and 1,211,694 and in GermanOffenlegungsschrift No. 1,800,698. Examples which may be mentioned aretri-lower alkyl-silyloxycarbonyl, such as trimethyl-silyloxycarbonyl andtert.-butyl-dimethyl-silyloxycarbonyl, lower alkoxy-loweralkyl-halogeno-silyloxycarbonyl, for examplechloro-methoxy-methyl-silyloxycarbonyl, or tri-loweralkyl-stannyloxycarbonyl, for example tri-n-butyl-stannyloxycarbonyl.

Esterified carboxyl groups R₂ ^(o) which are split easily by acidolysis,for example in the presence of hydrogen chloride, hydrogen fluoride orhydrogen bromide or of organic acids, such as acetic acid,trifluoroacetic acid or formic acid, if appropriate with the addition ofa nucleophilic compound, such as phenol or anisole, are derived fromlower alkanols which are poly-branched in the α-position or loweralkanols which contain one or more electron donors in the α-position.Examples of such esterified carboxyl groups are tert.butoxycarbonyl,tert.amyloxycarbonyl, cyclopentyloxycarbonyl, cyclohexyloxycarbonyl,adamantyloxycarbonyl, furfuryloxycarbonyl, p-methoxy-benzyloxycarbonyl,diphenylmethoxycarbonyl and pivaloyloxymethoxycarbonyl. Esterifiedcarboxyl groups R₂ ^(o) which can be split by reduction, for examplewith zinc and acid, are above all derived from 2-halogeno-loweralkanols, for example from 2,2,2-trichloroethanol and 2-iodoethanol.Carboxyl groups R₂ ^(o) esterified by phenacyl alcohol or p-nitrobenzylalcohol can be split by hydrogenolysis, for example by treatment withnascent hydrogen or with hydrogen in the presence of a noble metalcatalyst, for example a palladium catalyst.

Salts of starting materials in the formula II or IV are, in particular,salts of those compounds which have a free carboxyl group, above allammonium salts, such as tri-lower alkylammonium salts, for exampletriethylammonium salts, and also alkali metal salts.

The acylation of the amino group of the compound II with the carboxylicacid of the formula III is carried out in accordance with methods whichare in themselves known, in particular in the manner known frompenicillin chemistry and cephalosporin chemistry for the acylation ofweakly basic amino groups. The acylating agent used is either thecorresponding acid of the formula III, in which case the reaction iscarried out in the presence of a condensation agent, for example acarbodiimide, such as N,N'-diethyl-, N,N'-dipropyl-, N,N'-diisopropyl,N,N'-dicyclohexyl- or N-ethyl-N'-γ-dimethylamino-propylcarbodiimide, orin the presence of a suitable carbonyl compound, for exampleN,N'-carbonyldiimidazole, or of isoxazolium salts, for exampleN-ethyl-5-phenyl-isoxazolinium-3'-sulphonate orN-tert.butyl-5-methyl-isoxazolinium perchlorate, or of an acylaminocompound, for example 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline, oris a reactive functional derivative, above all an acid halide,especially an acid chloride or acid bromide, or, for example, anactivated ester, for example a p-nitrophenyl ester, 2,4-dinitrophenylester, 2,4,5- or 2,4,6-trichlorophenyl ester or pentachlorophenyl esteror, for example, the cyanomethyl ester, N-hydroxysuccinimide ester,N-hydroxypiperidine ester or N-hydroxyphthalimide ester, or a mixedanhydride, for example a mixed anhydride with mono-esterified carbonicacid, such as a carbonic acid lower alkyl ester, for example the ethylester or methyl ester, or a mixed anhydride with an optionallyhalogen-substituted lower alkanoic acid such as formic acid, pivalicacid or trichloroacetic acid. If a hydroxyl group is present in theα-position relative to the carboxyl group in the group B, a mixed inneranhydride having the partial formula ##STR15## can also be used for theacylation.

The acylation reaction is carried out in a solvent or diluent, ifdesired in the presence of a catalyst and/or in the presence of basicagents such as aliphatic, aromatic or heterocyclic nitrogen bases, forexample triethylamine, diisopropylethylamine, N,N-diethylaminoaceticacid ethyl ester, N-methylmorpholine, N,N-dimethylaniline, pyridine,2-hydroxypyridine, p-dimethylaminopyridine, collidine or 2,6-lutidine.

The solvents or diluents used are inert liquids, for example carboxylicacid amides, such as N,N-di-lower alkylamides, for exampledimethylformamide, or hexamethylphosphoric acid triamide, halogenatedhydrocarbons, for example methylene chloride, carbon tetrachloride orchlorobenzene, ketones, for example acetone, esters, for example ethylacetate, nitriles, for example acetonitrile, solvents containing oxagroups, such as tetrahydrofurane, and dioxane, or mixtures thereof.

The reaction is carried out at room temperature or with cooling orwarming, for example at temperatures of -70° to +100° C., if appropriatein an inert gas atmosphere, for example a nitrogen atmosphere, and/orwith exclusion of moisture.

In carrying out the acylation, free hydroxyl, mercapto, amino and/orcarboxyl groups which may be present in the reactants are advantageouslyprotected, especially by protective groups which can be split offeasily, such as are known, for example, from the synthesis of peptides,compare Schroder and Lubke "The Peptides", vol. 1, Academic Press, NewYork and London, 1965, and Th. Wieland, Angew, Chem. 63 (1951), 7-14, 66(1954), 507-512, 69 (1957), 362-372, 71 (1959), 417-425 and 75 (1963),539-551. Examples of amino protective groups which may be mentioned areoptionally substituted aralkyl groups, such as diphenylmethyl ortriphenylmethyl groups, or acyl groups, such as formyl, trifluoroacetyl,phthaloyl, p-toluenesulphonyl, benzylsulphonyl, benzenesulphenyl oro-nitrophenylsulphenyl, or, above all, groups derived from carbonic acidor thiocarbonic acid, such as carbobenzoxy groups optionally substitutedin the aromatic radical by halogen atoms, nitro groups or lower alkyl orlower alkoxy or lower carbalkoxy groups, for example carbobenzoxy,p-bromocarbobenzoxy or p-chlorocarbobenzoxy, p-nitrocarbobenzoxy orp-methoxycarbobenzoxy, coloured benzyloxycarbonyl groups, such asp-phenylazo-benzyloxycarbonyl andp-(p'-methoxyphenylazo)-benzyloxycarbonyl, tolyloxycarbonyl,2-phenyl-isopropoxycarbonyl, 2-tolyl-isopropoxycarbonyl and, above all,2-(para-biphenylyl)-2-propoxycarbonyl, and also aliphatic oxycarbonylgroups, such as, for example, allyloxycarbonyl, cyclopentyloxycarbonyl,tert.amyloxycarbonyl, adamantyloxycarbonyl,2,2,2-trichloroethoxycarbonyl, 2-iodoethoxycarbonyl and, above all,tert.butoxycarbonyl, and, for example, carbamoyl, thiocarbamoyl,N-phenylcarbamoyl and N-phenylthiocarbamoyl. Easily splittable estergroups which can be prepared to protect a free carboxyl group havealready been indicated above. Hydroxyl groups or mercapto groups can beprotected by etherification, for example with tert.butanol, or in theform of a trityl ether, a silyl ether, or a stannyl ether, or byesterification, for example with a half-ester of a carbonic acidhalf-halide, such as ethoxycarbonyl chloride.

The acylation of the amino group of the compound IV with the carboxylicacid of the formula V is carried out analogously.

In a compound obtained according to the invention, hydroxyl, mercapto,amino and carboxyl protective groups can be split off in a manner whichis in itself known by solvolysis, for example hydrolysis, alcoholysis oracidolysis, or by reduction, for example by hyrogenolysis.

In resulting compounds of the formula I, a substituted methyl group canbe converted into another group of this type. Thus, for example, acompound containing an esterified hydroxymethyl radical R₃, wherein theesterified hydroxyl group in particular denotes lower alkanoyloxy, forexample acetoxy, can first be reacted with thiobenzoic acid and then betreated with pyridine in the presence of a mercury salt, or can bereacted with a suitable salt, such as potassium thiocyanate, potassiumiodide or potassium nitrate, and with pyridine in the presence of waterat a pH value of about 6.5, obtained, for example, by means ofphosphoric acid, thus giving the corresponding pyridiniummethyl compoundwhich can, if required, be converted to the inner salt (the zwitter ionform), for example by treatment with a suitable ion exchange reagent.The pyridinium compound can also be manufactured in accordance with theprocess of Belgian Pat. No. 719,711 (DOS No. 1,795,643) by firstconverting the acetoxy group into a group more suitable for nucleophilicexchange, for example a halogen atom or an acetoxy group which containsan electron-attracting substituent, such as, for example, chloroacetoxy,dichloroacetoxy or cyanoacetoxy. Furthermore, compounds containing alower alkanoyloxymethyl group, for example an acetoxymethyl group, asthe radical R₃ can be reacted with a mercapto compound, such as anoptionally substituted lower alkylmercaptan, phenylmercaptan orheterocyclylmercaptan, thus giving compounds of the formula I, whereinR₃ represents an etherified mercaptomethyl group. The reaction iscarried out in water or in a mixture of water and a water-misciblesolvent, such as acetone, dioxane, dimethylformamide, tetrahydrofurane,hexamethylphosphoric acid triamide and the like, in the presence of abase, such as an alkali metal bicarbonate, for example sodiumbicarbonate, at a pH value of about 7 to 7.8, at temperatures from about10° C. to about 100° C., preferably at about 50° C. to about 60° C., andin an inert gas atmosphere, such as a nitrogen atmosphere.

Salts of compounds of the formula I can be manufactured in a mannerwhich is in itself known. Thus, salts of compounds of the formula I,wherein R₂ represents a free carboxyl group, can be formed, for example,by treatment with metal compounds, such as alkali metal salts ofsuitable carboxylic acids, for example with the sodium salt ofα-ethylcaproic acid, or with ammonia or a suitable organic amine.

Salts can be converted into the free compounds in the usual manner,metal salts and ammonium salts, for example, by treatment with suitableacids or ion exchangers.

Resulting mixtures of isomers can be separated into the individualisomers according to methods which are in themselves known, for exampleby fractional crystallisation, adsorption chromatography (columnchromatography or thin layer chromatography) or other suitable methodsof separation. Resulting racemates can be separated into the antipodesin the usual manner, if appropriate after the introduction of suitablesalt-forming groupings, for example by forming a mixture ofdiastereoisomeric salts with optically active salt-forming agents,separating the mixture into the diastereoisomeric salts and convertingthe separated salts into the free compounds, or by fractionalcrystallisation from optically active solvents.

The process also embraces those embodiments according to which compoundsobtained as intermediates are used as starting materials and theremaining process steps are carried out with these, or the process isdiscontinued at any stage; furthermore, starting materials can be usedin the form of derivatives or be formed during the reaction.

Preferably, those starting materials are used, and the reactionconditions are so chosen, that the compounds mentioned initially asbeing particularly preferred are obtained.

The starting materials of the formula II, III and IV are known or can bemanufactured according to known processes.

Compounds of the formula V and their reactive functional derivativeshave not previously been disclosed. They can be obtained in a mannerwhich is in itself known, by acylating a compound of the formula R₁--CH(NH₂)--COOH (VI), in which the carboxyl group is present in aprotected form, for example in the form of an easily splittableesterified carboxyl group, with a carboxylic acid of the formula B-COOH(III), with a halide thereof or with a mixed anhydride thereof, thensplitting off the carboxyl protective group and, if desired, convertingthe carboxylic acid obtained into a reactive functional derivativethereof.

Easily splittable esters of carboxylic acids of the formula VI are, inparticular, those which after acylation of the amino group can be splitsolvolytically, for example hydrolytically, alcoholytically orespecially acidolytically, or reductively, to give the free carboxylicacid of the formula V. Such easily splittable ester groups are derivedfrom the same alcohols from which the easily splittable esterifiedcarboxyl groups R₂ ^(o) are derived, and the subsequent splitting iscarried out analogously. Benzyl esters of compounds of the formula VIwhich, after N-acylation, can be split reductively, for example by meansof palladium and hydrogen, are preferred.

The acylation of the amino group in a compound of the formula (VI), inwhich the carboxyl group is protected, is carried out analogously to theacylation of the amino group in a compound of the formula (II), it beingpossible to use the same halides, mixed anhydrides or inner mixedanhydrides having the partial formula (IIIa), of the carboxylic acid ofthe formula III, or this carboxylic acid itself.

The conversion of a resulting carboxylic acid of the formula (V) into areactive functional derivative thereof is carried out in a manner whichis in itself known. Carboxylic acid chlorides are obtained, for example,by treatment with thionyl chloride, activated esters are obtained, forexample, by reaction of this resulting carboxylic acid chloride with anappropriate hydroxy compound, for example p-nitrophenol orN-hydroxyphthalimide, and mixed anhydrides are obtained by reaction of acarboxylic acid of the formula (V) with an appropriate halide, forexample chloride, of a second carboxylic acid, for example amono-esterified carbonic acid, such as a carbonic acid lower alkylester, for example the ethyl ester or methyl ester, or an optionallyhalogen-substituted lower alkanoic acid, such as formic acid, pivalicacid or trichloroacetic acid. When forming the mixed anhydride, asimultaneously present hydroxyl or mercapto group in the radical B can,if appropriate, be esterified by the carboxylic acid halide used, forexample by the ethoxycarbonyl radical when using chloroformic acid ethylester.

The new compounds can be used as medicaments, for example in the form ofpharmaceutical preparations which contain an effective amount of theactive substance together with, or mixed with, inorganic or organic,solid or liquid, pharmaceutically usable excipients which are suitablefor enteral or, preferably, parenteral administration. Thus, tablets orgelatine capsules are used, which contain the active compound togetherwith excipients, for example lactose, sucrose, mannitol, sorbitol,cellulose and/or glycine, and lubricants, for example silica, talc,stearic acid or salts thereof, such as magnesium stearate or calciumstearate, and/or polyethylene glycol; tablets also contain binders, forexample magnesium aluminium silicate, starches, such as maize starch,wheat starch, rice starch or arrowroot, gelatine, tragacanth,methylcellulose, sodium carboxymethylcellulose and/orpolyvinylpyrrolidone and, if desired, disintegrating agents, for examplestarches, agar, alginic acid or a salt thereof, such as sodium alginate,and/or effervescent mixtures, or adsorbents, dyestuffs, flavouringsubstances and sweeteners. Preferably, the pharmacologically activecompounds of the present invention are used in the form of injectablepreparations, for example intraveneously administrable preparations, orof infusion solutions. Such solutions are, preferably, isotonic aqueoussolutions or suspensions and these can be prepared before use, forexample from lyophilised preparations which contain the active compoundby itself or together with an excipient, for example mannitol. Thepharmacological preparations can be sterilised and/or containauxiliaries, for example preservatives, stabilisers, wetting agentsand/or emulsifiers, solubilising agents, salts for regulating theosmotic pressure and/or buffers. The present pharmaceutical preparationswhich, if desired, can contain further pharmacologically valuablematerials, are produced in a manner which is in itself known, forexample by means of conventional mixing, granulating, dragee-making,dissolving or lyophilising processes, and contain from about 0.1% to100%, especially from about 1% to about 50%, of lyophilisates up to 100%of the active compound.

In the context of the present description, organic radicals described as"lower" contain up to 7, preferably up to 4, carbon atoms; acyl radicalscontain up to 20, preferably up to 12, carbon atoms.

The examples which follow serve to illustrate the invention.

The following systems are used in the thin layer chromatography:

System 52 A: n-butanol/glacial acetic acid/water (67:10:23)

System 67: n-butanol/ethanol/water (40:10:50, upper phase)

System 101: n-butanol/pyridine/glacial acetic acid/water (38:24:8:30)

System 101A: n-butanol/pyridine/glacial acetic acid/water (42:24:4:30).

EXAMPLE 1

22.6 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 32.4 g ofD(-)-cephaloglycine in 300 ml of methylene chloride, a clear solutionbeing formed. This mixture is then cooled to -10° C. and a solution of12.0 g of 2-hydroxy-pyridine-5-carboxylic acid chloride in a mixture of50 ml of N,N-dimethylformamide and 50 ml of methylene chloride is addeddropwise at -10° C. in the course of 30 minutes, whilst stirring andcooling. The suspension is then diluted with 200 ml of methylenechloride, stirred for 1 hour at 0° C., then treated with 200 ml of waterand with 1 N aqueous sodium bicarbonate solution until a pH value of 7.2is obtained and mixed vigorously. The methylene chloride phase isseparated off and the aqueous phase is extracted twice with ethylacetate, then cooled in an ice bath and acidified (pH 2.5) with 1 Nhydrochloric acid, whilst stirring. The product obtained is filtered offand washed successively with ethanol, methanol and ether. The materialon the filter is suspended in 200 ml of water and the solid product isbrought into solution by the addition of 2 N aqueous sodium bicarbonatesolution (pH 7.2) and the insoluble by-product (a small amount) isfiltered off. The filtrate is cooled in an ice bath and acidified (pH2.5) with 1 N hydrochloric acid. The product obtained is filtered off,washed with a large amount of cold water and dried at 30° C./0.1 mm Hg.7β-[D(-)-α-(2-Hydroxy-pyridine-5-carboxamido)-phenylacetamido]-cephalosporanicacid melts at 242°-245° C. with decomposition. Thin layer chromatogramon silica gel: Rf_(52A) =0.28, Rf₆₇ =0.23, Rf₁₀₁ =0.50, Rf_(101A) =0.42,[α]_(D) ²⁰ =+27°±1° (c=1.063 in dimethylsulphoxide).

EXAMPLE 2

25.0 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 14.6 g ofD(-)-cephaloglycine in 300 ml of methylene chloride, a clear solutionbeing formed. This mixture is then cooled to -10° C. and a solution of5.6 g of 1,6-dihydro-6-oxo-3-pyridazinecarboxylic acid chloride in 60 mlof N,N-dimethylformamide is added dropwise at -10° C. in the course of30 minutes, whilst stirring and cooling. The suspension is then stirredfor 1 hour at 0° C. and for 1 hour at room temperature. The clearsolution is then concentrated to half its volume on a rotary evaporatorat 40° C. and the product is precipitated by adding 500 ml of ether andfiltered off. The material on the filter is dissolved in 100 ml ofwater, 2 N aqueous sodium bicarbonate solution being added until a pHvalue of 7.2 is reached. The solution is filtered through celite and thefiltrate is covered with ethyl acetate, rendered acid (pH 2.5) at 10°C., whilst stirring and cooling in an ice bath, by adding 20% strengthphosphoric acid and extracted three times with ethyl acetate. The ethylacetate extracts are combined, washed with twice 50 ml of sodiumchloride solution, dried over sodium sulphate and freed from solvent ona rotary evaporator at 45° C. The residual7β-[D(-)-α-(1,6-dihydro-6-oxo-3-pyridazinecarboxamido)-phenylacetamido]-cephalosporanicacid is purified by crystallising from methanol. Melting point:210°-212° C. with decomposition. Thin layer chromatogram on silica gel:Rf_(52A) =0.28, Rf₆₇ =0.24, Rf₁₀₁ =0.52, Rf_(101A) =0.45; [α]_(D) ²⁰=+48°±1 (c=0.798, in dimethylsulphoxide).

EXAMPLE 3

2.8 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 4.0 g ofD(-)-cephaloglycine in a mixture of 20 ml of methylene chloride and 20ml of tetrahydrofurane, a clear solution being formed. This mixture isthen cooled to -10° C. and a suspension of 2.2 g of2-hydroxy-quinoline-4-carboxylic acid chloride in a mixture of 10 ml ofN,N-dimethylformamide and 10 ml of methylene chloride is added dropwiseat -10° C. in the course of 20 minutes, whilst stirring and cooling. Thereaction mixture is diluted with 15 ml of tetrahydrofurane and stirredfor 1.5 hours at 0° C. The product is precipitated by adding 100 ml ofether and filtered off. The material on the filter is dissolved in 50 mlof a phosphate buffer solution of pH 7.5 and the solution is extractedtwice with ethyl acetate. The aqueous phase is separated off, coveredwith ethyl acetate, rendered acid (pH 2.5) at 10° C., whilst stirringand cooling in an ice bath, by adding 20% strength phosphoric acid andextracted three times with ethyl acetate. The ethyl acetate extracts arecombined, washed twice with sodium chloride solution, dried over sodiumsulphate and freed from solvent on a rotary evaporator at 45° C. Theresidual product is purified by crystallising from methanol.7β-[D(-)-α-(2-Hydroxyquinoline-4-carboxamido)-phenylacetamido]-cephalosporanicacid melts at 218°-221° C. with decomposition. Thin layer chromatogramon silica gel: Rf_(52A) =0.35, Rf₆₇ =0.32, Rf₁₀₁ =0.57, Rf_(101A) =0.48;[α]_(D) ²⁰ =+27°±1° (c= 1.008, in dimethylsulphoxide).

EXAMPLE 4

4.20 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 4.05 g ofD(-)-cephaloglycine in 30 ml of methylene chloride, a clear solutionbeing formed. This mixture is then cooled to -10° C. and a solution of2.31 g of 2,6-dichloro-pyridine-4-carboxylic acid chloride in 20 ml ofmethylene chloride is added dropwise at -10° C. in the course of 15minutes, whilst stirring and cooling. Thereafter, the suspension isstirred for 1.5 hours at -10° C., then diluted with 200 ml of ethylacetate and extracted with 100 ml of a phosphate buffer solution of pH7.5. The aqueous phase is separated off, extracted twice with ethylacetate (neutral extract), then covered with ethyl acetate, renderedacid (pH 2.5) at 10° C., whilst stirring and cooling in an ice bath, byadding 20% strength phosphoric acid and extracted three times with ethylacetate. The last three ethyl acetate extracts are combined, washedtwice with sodium chloride solution and dried over sodium sulphate andthe solvent is evaporated on a rotary evaporator at 45° C. The residualproduct is purified by crystallising from methanol.7β-[D(-)-α-(2,6-Dichloro-pyridine-4-carboxamido-phenylacetamido]-cephalosporanicacid melts at 193°-195° C. with decomposition. Thin layer chromatogramon silica gel: Rf_(52A) =0.44, Rf₆₇ =0.50, Rf₁₀₁ =0.59, Rf_(101A) =0.55.[α]_(D) ²⁰ =+26°±1° (c=1.063 in dimethylsulphoxide).

EXAMPLE 5

21.0 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 20.0 g ofD(-)-cephaloglycine in a mixture of 300 ml of methylene chloride and 600ml of tetrahydrofurane, a clear solution being formed. This mixture isthen cooled to 0° C. and a solution of 11.0 g of3-methylmercapto-5-hydroxy-1,2,4-triazine-6-carboxylic acid chloride ina mixture of 40 ml of N,N-dimethylformamide and 40 ml oftetrahydrofurane is added dropwise at 0° C. in the course of 30 minutes,whilst stirring and cooling. The suspension is then stirred for 1 hourat room temperature and then concentrated to a volume of about 100 ml ona rotary evaporator (waterpump vacuum) at 45° C. The product isprecipitated by adding 400 ml of ether, filtered off and washed withwater. The material on the filter is worked up as in Example 3. Afterrecrystallisation from ethyl acetate,7β-[D(-)-α-(3-methylmercapto-5-hydroxy-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid melts at 235°-240° C. with decomposition. Thin layer chromatogramon silica gel: Rf_(52A) =0.28, Rf₆₇ =0.25, Rf₁₀₁ =0.50, Rf_(101A) =0.41.[α]_(D) ²⁰ =+42°±1° (c=0.994 in dimethylsulphoxide).

EXAMPLE 6

13.0 ml of triethylamine are added, at room temperature, whilst stirringand with exclusion of atmospheric moisture, to a suspension of 12.5 g ofD(-)-cephaloglycine in a mixture of 200 ml of methylene chloride and 300ml of tetrahydrofurane, a clear solution being formed. This mixture isthen cooled to 0° C. and a solution of 7.65 g of3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxylic acid chloridein a mixture of 40 ml of N,N-dimethylformamide and 40 ml oftetrahydrofurane is added dropwise at 0° C. in the course of 30 minutes,whilst stirring and cooling. The suspension is then stirred for 1 hourat room temperature and then concentrated to a volume of about 100 ml ona rotary evaporator (waterpump vacuum) at 45° C. The product isprecipitated by adding 400 ml of ether, filtered off and washed withether. The material on the filter is worked up as in Example 3.7β-[D(-)-α-(3,5-Dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid crystallises from ethyl acetate. Melting point: 153°-158° C. withdecomposition. Thin layer chromatogram on silica gel: Rf_(52A) =0.32,Rf₆₇ =0.20, Rf₁₀₁ =0.52, Rf_(101A) =0.48. [α]_(D) ²⁰ =+31°±1° (c=1.089in dimethylsulphoxide).

EXAMPLE 7

5.88 ml of triethylamine are added, at room temperature whilst stirringand with exclusion of atmospheric moisture, to a suspension of 2.55 g of7β-[D(-)-α-amino-phenylacetamido]-3-methoxy-ceph-3-em-4-carboxylic aciddihydrate in 200 ml of absolute dimethylformamide, a clear solutionbeing formed. This mixture is then cooled to 0° C. and a solution of1.85 g of 3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxylic acidchloride in a mixture of 7 ml of N,N-dimethylformamide and 12 ml oftetrahydrofurane is added dropwise at 0° C. in the course of 10 minutes,whilst stirring and cooling. The suspension is then stirred for 30minutes at 0° C. and for 2 hours at room temperature. The product isprecipitated by adding 2 l of ether, filtered off and washed with ether.The material on the filter is dissolved in 100 ml of a phosphate buffersolution of pH 7.5 and the solution is extracted twice with ethylacetate. The aqueous phase is separated off, covered with ethyl acetate,rendered acid (pH 2.5) at 10° C., whilst stirring and cooling in an icebath, by adding 20% strength phosphoric acid and extracted three timeswith ethyl acetate. The ethyl acetate extracts are combined, washedthree times with sodium chloride solution and dried over sodium sulphateand the solvent is evaporated on a rotary evaporator. The residualproduct is purified by dissolving in acetone and precipitating withethyl acetate.7β-[D(-)-α-(3,5-Dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-methoxy-ceph-3-em-4-carboxylicacid melts at 183°-185° C. with decomposition. Thin layer chromatogramon silica gel: Rf_(52A) : 0.33, [α]_(D) =+78°±1° (c=1.0, methanol).

EXAMPLE 8

A solution of 1.90 g of 2-amino-5-mercapto-1,3,4-thiadiazole (preparedaccording to the method of J. Sandstrom, Acta chem. Scand. 15, 1295(1961)) in 20 ml of 0.5 N aqueous sodium bicarbonate solution is addedto a clear solution of 7.05 g of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid (prepared as in Example 6) in a mixture of 30 ml of water and 70 mlof 0.5 N aqueous sodium bicarbonate solution and the reaction mixture isstirred for 20 hours at 60° C. in a nitrogen atmosphere. At the start ofthe reaction the pH value of the reaction mixture rises and is broughtback to a pH value of 7.5 to 7.6 by the occasional dropwise addition of0.1 N hydrochloric acid. After the reaction time has elapsed, thereaction mixture is cooled to room temperature and extracted twice withethyl acetate. The aqueous phase is separated off, cooled in an ice bathand acidified (pH 3.0) with 20% strength phosphoric acid and the solidproduct obtained is filtered off and washed with water. The material onthe filter is then suspended in 1 liter of ethyl acetate and thesuspension is stirred for 5 minutes at room temperature and filtrated.The material on the filter is then introduced into 1 liter of a mixtureof methanol/tetrahydrofurane (1:1) and the mixture is stirred for 10minutes at room temperature. The insoluble fraction is filtered off andthe filtrate is evaporated on a rotary evaporator (waterpump vacuum) at45° C. The residual7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)phenylacetamido]-3-[(2-amino-1,3,4-thiadiazol-5-ylthio)methyl]-ceph-3-em-4-carboxylicacid crystallises on mixing with ethyl acetate. Melting point: 255°-260°C. with decomposition. Thin layer chromatogram on silica gel: Rf_(52A)=0.32, Rf₆₇ =0.20, Rf₁₀₁ =0.52, Rf_(101A) =0.48. [α]_(D) ²⁰ =-64°±1°(c=1.036 in dimethylsulphoxide).

EXAMPLE 9

A solution of 0.93 g of2-(β-carboxy-propionylamido)-5-mercapto-1,3,4-thiadiazole in 11 ml of0.5 N aqueous sodium bicarbonate solution is added to a clear solutionof 2.0 g of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid (prepared as in Example 6) in a mixture of 10 ml of water and 20 mlof 0.5 N aqueous sodium bicarbonate solution and the reaction mixture isstirred for 20 hours at 60° C. in a nitrogen atmosphere. At the start ofthe reaction the pH value of the reaction mixture rises and is broughtback to a pH value of 7.5 to 7.6 by the occasional dropwise addition of0.1 N hydrochloric acid. After the reaction time has elapsed, thereaction mixture is cooled to room temperature and extracted twice withethyl acetate. The aqueous phase is separated off, cooled in an ice bathand acidified (pH 2.5) with 20% strength phosphoric acid and the solidproduct obtained is filtered off and washed with water. The material onthe filter is dissolved in a mixture of 20 ml of methanol and 20 ml oftetrahydrofurane and the solution is concentrated to a volume of about10 ml on a rotary evaporator (waterpump vacuum) at 45° C. and ethylacetate is then added, whereupon the product crystallises. Afterrecrystallisation from ethyl acetate,7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(2-(β-carboxy-propionylamido)-1,3,4-thiadiazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid melts at 226°-227° C. with decomposition. Thin layer chromatogramon silica gel: Rf₅₂ =0.26, Rf₆₇ =0.05, Rf_(101A) =0.36. [α]_(D) ²⁰=-50°±1° (c-1.234 in dimethylsulphoxide).

Preparation of the starting material

23 ml of triethylamine are added to a solution of 20.0 g of2-amino-5-mercapto-1,3,4-thiadiazole (prepared according to the methodof J. Sandstrom, Acta Chem. Scand. 15, 1295 (1961)) in 600 ml ofdioxane, a suspension being formed. This suspension is cooled to 10° C.using an ice bath, and 25.0 g of succinic acid monomethyl ester chlorideare added dropwise at 10°-13° C. in the course of 30 minutes, whilststirring vigorously, and the reaction mixture is then stirred for 20hours at room temperature. The precipitate is then filtered off, thematerial on the filter is washed with a little dioxane and the filtrateis evaporated to dryness on a rotary evaporator (high vacuum) at 40° C.The residual solid product is mixed with a little water and filteredoff. 28.0 g of the crude product are suspended in 840 ml of 1 N sodiumbicarbonate solution, the suspension is stirred vigorously for 5 minutesat room temperature, the insoluble fraction is filtered off and theaqueous filtrate is cooled to 3° C. in an ice bath and is rendered acid(pH 2.5) by the dropwise addition of 2 N hydrochloric acid. The productobtained is filtered off and washed with cold water. Afterrecrystallisation from hot water,2-(β-carbomethoxypropionylamido)-5-mercapto-1,3,4-thiadiazole melts at210°-212° C. with decomposition. Thin layer chromatogram on silica gel(running agent: chloroform/ethyl acetate/glacial acetic acid(66:33:0.25)): Rf=0.17.

A solution of 17.0 g of2-(β-carbomethoxypropionylamido)-5-mercapto-1,3,4-thiadiazole in amixture of 130 ml of water and 155 ml of 1 N aqueous sodium hydroxidesolution is stirred for 20 hours at room temperature. Thereafter theyellow solution is filtered and the filtrate is then cooled to 5° C. inan ice bath and rendered acid (pH 2.0) by the dropwise addition ofdilute hydrochloric acid (1:1). The product obtained is filtered off andwashed with cold water. After recrystallisation from hot water,2-(β-carboxypropionylamido)-5-mercapto-1,3,4-thiadiazole melts at225°-230° C. with decomposition. Thin layer chromatogram on silica gel(running agent: chloroform/ethyl acetate/glacial acetic acid(66:33:1.5)): Rf=0.15.

EXAMPLE 10

In accordance with the process of Example 8,7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(1-methyl-1H-tetrazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid is obtained by reacting 5.44 g of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid (prepared as in Example 6) with 1.30 g of5-mercapto-1-methyl-tetrazole in water in the presence of sodiumbicarbonate.

EXAMPLE 11

In accordance with the process of Example 7,7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(1H-1,2,3-triazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid is obtained by reacting 6.00 g of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid (prepared as in Example 6) with 1.25 g of5-mercapto-1H-1,2,3-triazole (prepared in accordance with the method ofJ. Goerdeler and G. Gnad, Chem. Ber. 99, 1618 (1966)) in water in thepresence of sodium bicarbonate.

EXAMPLE 12

6 ml of a 40% strength aqueous mercury perchlorate solution are added toa solution of 1.5 g of the sodium salt of3-benzoylthiomethyl-7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-ceph-3-em-4-carboxylicacid and 3.2 g of isonicotinic acid amide in 15 ml of water and 15 ml ofdioxane and the reaction mixture is stirred vigorously for 1 hour at 45°C. in a nitrogen atmosphere. The suspension is then cooled to 0° C., 4ml of thiobenzoic acid are added dropwise, the mixture is stirred for 5minutes at +10° C. and the mercury salt obtained is removed by filteringthrough celite and the material on the filter is washed with a 1:1mixture of water and dioxane. The filtrate is washed successively twicewith petroleum ether, twice with, in each case, 100 ml of a 10% strengthsolution of "Amberlite" LA-2 in petroleum ether and finally withpetroleum ether. The aqueous phase is separated off and evaporated todryness on a rotary evaporator (high vacuum) at 45° C. The solid residueis mixed well with a mixture of methanol/ethanol/diethyl ether (1:1:1)and the insoluble7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[1-(4-carbamoyl-pyridiniomethyl)]-ceph-3-em-4-carboxylateis filtered off and washed with the above solvent mixture.

Preparation of the starting material

A solution of 10.0 g of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid (prepared as in Example 6) in 20 ml of water and 46 ml of 1 Nsodium bicarbonate solution is added to a solution of 9.0 g ofthiobenzoic acid and 6.05 g of solid sodium bicarbonate in 50 ml ofwater and the reaction mixture is stirred for 20 hours at 50°-55° C. ina nitrogen atmosphere. At the start of the reaction the pH value of thereaction mixture rises and is brought back to a pH value of 7.6 to 7.7by the occasional dropwise addition of 0.1 N hydrochloric acid. Afterthe reaction time has elapsed, the suspension is cooled to 10° C. andthe sodium salt of3-benzoylthiomethyl-7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-ceph-3-em-4-carboxylicacid, which is obtained, is filtered off and washed with a little coldwater. Melting point: 215°-220° C. with decomposition. Thin layerchromatogram on silica gel: Rf_(52A) =0.41; Rf_(101A) =0.53.

EXAMPLE 13

Dry powders or phials, containing 0.6 g of the sodium salt of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid are prepared as follows:

Composition (for 1 ampoule or phial)

Sodium salt of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]cephalosporanicacid: 0.6 g

Mannitol: 0.06 g

A sterile aqueous solution of the sodium salt of7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-cephalosporanicacid and of the mannitol is subjected to freeze-drying in 5 ml ampoulesor 5 ml phials under aseptic conditions and the ampoules or phials aresealed and tested.

We claim:
 1. A compound of the formula ##STR16## wherein R₁ denotesphenyl, thienyl, furyl, 1,4-cyclohexadienyl, or hydroxy phenyl, R₂represents a free carboxyl group, R₃ represents heterocyclylthiomethyl,wherein heterocyclyl has 5 ring members is of aromatic character,comprises as hetero atoms 2 nitrogen atoms and a further oxygen orsulphur atom or 1 to 2 further nitrogen atoms, such a heterocyclus beingunsubstituted or substituted by lower alkyl with 1-5 carbon atoms, loweralkoxy or lower alkylthio with 1-5 carbon atoms, cycloalkyl with 3-7carbon atoms, phenyl, or phenyl substituted by a nitro group, by loweralkyl with 1 to 5 carbon atoms, lower alkoxy with 1 to 5 carbon atoms ora halogen, thienyl, or thienyl substituted as indicated for phenyl,amino, lower alkylamino, or lower alkylamino substituted in the alkylgroup by carboxyl or amino, lower alkanoylamino, or lower alkanoylaminosubstituted on the alkyl by carboxyl or amino, lower alkoxycarbonylamino or sulphonylamino, and B represents one of the groups B₅or B₆, of which B₅ represents the formula ##STR17## wherein R₁₄ and R₁₅independently of one another denote hydrogen, halogen, a free hydroxylor mercapto group, a lower alkoxy or a lower alkylmercapto group or alower alkoxycarbonyloxy or a lower alkoxycarbonylmercapto group, ormono- or di-lower alkylated amino group, a lower alkanoylated aminogroup, or ureido group or of which B₆ represents the formula ##STR18##wherein R₁₄ and R₁₅ have the meaning mentioned under formula (B₅), thesubstituent R₁₄ preferably occupying the 5-position of the as-triazinering, a tautomer, or a pharmaceutically acceptable salt thereof.
 2. Acompound of the formula I according to claim 1, wherein R₁ and R₂ and R₃have the meaning mentioned under formula I and B is 1,2,4-triazine,di-substituted by hydroxyl and/or methylmercapto, or a tautomer thereof,or a salt thereof.
 3. A compound of formula I according to claim 1,wherein R₁ represents phenyl, R₂ represents carboxyl, R₃ represents1,3,4-thiadiazolylthiomethyl, 1,2,3-triazolylthiomethyl ortetrazolylthiomethyl, which groups are unsubstituted or substituted asindicated in claim 14, and wherein B represents 1,2,4-triazinedisubstituted by hydroxyl and/or methylmercapto, or a tautomer thereofor a salt thereof.
 4. Pharmaceutical preparations for combattingbacteria which comprise an antibacterially effective amount of acompound of claim 1 or a pharmaceutically active salt thereof and apharmaceutically usable excipient.
 5. A compound according to claim 1which is7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(2-amino-1,3,4-thiadiazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid or a pharmaceutically acceptable salt thereof.
 6. A compoundaccording to claim 1 which is7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(2-(β-carboxypropionylamido)-1,3,4-thiadiazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid or a pharmaceutically acceptable salt thereof.
 7. Thepharmaceutical preparation of claim 4 wherein the active compound is7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(2-amino-1,3,4-thiadiazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid or a pharmaceutically acceptable salt thereof.
 8. Thepharmaceutical preparation of claim 4 wherein the active compound is7β-[D(-)-α-(3,5-dioxo-2,3,4,5-tetrahydro-1,2,4-triazine-6-carboxamido)-phenylacetamido]-3-[(2-β-carboxypropionylamido)-1,3,4-thiadiazol-5-ylthio)-methyl]-ceph-3-em-4-carboxylicacid or a pharmaceutically acceptable salt thereof.